A Consecutive Genome Engineering Method Reveals a New Phenotype and Regulation of Glucose and Glycerol Utilization in .

is a microorganism for production of 1,3-propanediol (1,3-PDO) and butanol, but suffers from lacking genetic tools for metabolic engineering to improve product titers. Furthermore, previous studies of have mainly focused on single genomic modification. The aim of this work is the development and application of a method for modification of multiple gene targets in the genome of .

Lactate-mediated mixotrophic co-cultivation of Clostridium drakei and recombinant Acetobacterium woodii for autotrophic production of volatile fatty acids.

Background: Acetogens, a diverse group of anaerobic autotrophic bacteria, are promising whole-cell biocatalysts that fix CO during their growth. However, because of energetic constraints, acetogens exhibit slow growth and the product spectrum is often limited to acetate. Enabling acetogens to form more valuable products such as volatile fatty acids during autotrophic growth is imperative for cementing their place in the future carbon neutral industry.

Biodegradable Silk Fibroin Matrices for Wound Closure in a Human 3D Ex Vivo Approach.

In this study, the potential of silk fibroin biomaterials for enhancing wound healing is explored, focusing on their integration into a human 3D ex vivo wound model derived from abdominoplasties. For this purpose, cast silk fibroin membranes and electrospun nonwoven matrices from silk cocoons were compared to untreated controls over 20 days. Keratinocyte behavior and wound healing were analyzed qualitatively and quantitatively by histomorphometric and immune histochemical methods (HE, Ki67, TUNEL).

Implementation of mDoE-methods to a microcarrier-based expansion processes for mesenchymal stem cells.

The need for advanced therapy medicinal products (ATMPs) has gained increased attention in recent years. In this respect, a well-designed cell expansion process is needed to efficiently manufacture the required number of cells with the desired product quality. This step is challenging due to the biological complexity of the respective primary cell (e.

Deep-learning-assisted Sort-Seq enables high-throughput profiling of gene expression characteristics with high precision.

Owing to the nondeterministic and nonlinear nature of gene expression, the steady-state intracellular protein abundance of a clonal population forms a distribution. The characteristics of this distribution, including expression strength and noise, are closely related to cellular behavior. However, quantitative description of these characteristics has so far relied on arrayed methods, which are time-consuming and labor-intensive.

Design, Evaluation, and Implementation of Synthetic Isopentyldiol Pathways in .

Isopentyldiol (IPDO) is an important raw material in the cosmetic industry. So far, IPDO is exclusively produced through chemical synthesis. Growing interest in natural personal care products has inspired the quest to develop a biobased process.

Electrochemical HO - stat mode as reaction concept to improve the process performance of an unspecific peroxygenase.

The electroenzymatic hydroxylation of 4-ethylbenzoic acid catalyzed by the recombinant unspecific peroxygenase from the fungus Agrocybe aegerita (rAaeUPO) was performed in a gas diffusion electrode (GDE)-based system. Enzyme stability and productivity are significantly affected by the way the co-substrate hydrogen peroxide (HO) is supplied. In this study, two in-situ electrogeneration modes of HO were established and compared.

Lactate based caproate production with and process control of via lactate dependent electrolysis.

Syngas fermentation processes with acetogens represent a promising process for the reduction of CO emissions alongside bulk chemical production. However, to fully realize this potential the thermodynamic limits of acetogens need to be considered when designing a fermentation process. An adjustable supply of H as electron donor plays a key role in autotrophic product formation.

Electrochemically mediated bioconversion and integrated purification greatly enhanced co-production of 1,3-propanediol and organic acids from glycerol in an industrial bioprocess.

In this study, we show how electrochemically mediated bioconversion can greatly increase the co-production of 1,3-propanediol and organic acids from glycerol in an industrial bioprocess using a Clostridum pasteurianum mutant. Remarkably, an enhanced butyrate formation was observed due to a weakened butanol pathway of the mutant. This allowed the strain to have a higher ATP generation for an enhanced growth, higher glycerol consumption and PDO production.

Novel synthetic co-culture of and using CO and in situ generated H for the production of caproic acid via lactic acid.

is known to produce mainly acetate from CO and H, but the production of higher value chemicals is desired for the bioeconomy. Using chain-elongating bacteria, synthetic co-cultures have the potential to produce longer-chained products such as caproic acid. In this study, we present first results for a successful autotrophic co-cultivation of mutants and a wild-type strain in a stirred-tank bioreactor for the production of caproic acid from CO and H via the intermediate lactic acid.

Comparison of mechanistic and hybrid modeling approaches for characterization of a CHO cultivation process: Requirements, pitfalls and solution paths.

Despite the advantages of mathematical bioprocess modeling, successful model implementation already starts with experimental planning and accordingly can fail at this early stage. For this study, two different modeling approaches (mechanistic and hybrid) based on a four-dimensional antibody-producing CHO fed-batch process are compared. Overall, 33 experiments are performed in the fractional factorial four-dimensional design space and separated into four different complex data partitions subsequently used for model comparison and evaluation.

Generation and evaluation of input values for computational analysis of transport processes within tissue cultures.

Techniques for tissue culture have seen significant advances during the last decades and novel 3D cell culture systems have become available. To control their high complexity, experimental techniques and their Digital Twins (modelling and computational tools) are combined to link different variables to process conditions and critical process parameters. This allows a rapid evaluation of the expected product quality.

Control of redox potential in a novel continuous bioelectrochemical system led to remarkable metabolic and energetic responses of Clostridium pasteurianum grown on glycerol.

Background: Electro-fermentation (EF) is an emerging tool for bioprocess intensification. Benefits are especially expected for bioprocesses in which the cells are enabled to exchange electrons with electrode surfaces directly. It has also been demonstrated that the use of electrical energy in BES can increase bioprocess performance by indirect secondary effects.

Food web and metabolic interactions of the lung inhabitants Streptococcus pneumoniae and Pseudomonas aeruginosa.

Bacteria that successfully adapt to different substrates and environmental niches within the lung and overcome the immune defence can cause serious lung infections. Such infections are generally complex, and recognized as polymicrobial in nature. Both Pseudomonas aeruginosa and Streptococcus pneumoniae can cause chronic lung infections and were both detected in cystic fibrosis (CF) lung at different stages.

Enhanced sequestration of carbon dioxide into calcium carbonate using pressure and a carbonic anhydrase from alkaliphilic .

CO in the atmosphere is a major contributor to global warming but at the same time it has the potential to be a carbon source for advanced biomanufacturing. To utilize CO, carbonic anhydrase has been identified as a key enzyme. Furthermore, attempts have been made to accelerate the sequestration via pressure.

Biosynthesizing structurally diverse diols via a general route combining oxidative and reductive formations of OH-groups.

Diols encompass important bulk and fine chemicals for the chemical, pharmaceutical and cosmetic industries. During the past decades, biological production of C3-C5 diols from renewable feedstocks has received great interest. Here, we elaborate a general principle for effectively synthesizing structurally diverse diols by expanding amino acid metabolism.

Biosynthesis Based on One-Carbon Mixotrophy.

Recent advances in biosynthesis using one-carbon (C1) compounds (e.g., CO and syngas) have led to first process examples of industrial demonstration for producing C1-based chemicals.

Understanding and Engineering Glycine Cleavage System and Related Metabolic Pathways for C1-Based Biosynthesis.

The glycine cleavage system (GCS) is a fundamental component of life, widely existing in microbes, plants, animals, and humans. A better understanding of the functionality and working mechanisms, and the engineering of the GCS have both scientific and practical impacts, which may lead to new knowledge and findings in life sciences, improved biomass production and human/animal health, efficient biosynthesis of chemicals, effective carbon fixation and global climate change mitigation. In this chapter, the GCS is first discussed in the context of the reductive glycine pathway (rGlyP), a recently proposed and appealing assimilation pathway of CO and formate, and its implementation and optimization in microorganisms for formatotrophic growth.

Strain evolution and novel downstream processing with integrated catalysis enable highly efficient coproduction of 1,3-propanediol and organic acid esters from crude glycerol.

Bioconversion of natural microorganisms generally results in a mixture of various compounds. Downstream processing (DSP) which only targets a single product often lacks economic competitiveness due to incomplete use of raw material and high cost of waste treatment for by-products. Here, we show with the efficient microbial conversion of crude glycerol by an artificially evolved strain and how a catalytic conversion strategy can improve the total products yield and process economy of the DSP.

Phenotype analysis of cultivation processes via unsupervised machine learning: Demonstration for .

A novel approach of phenotype analysis of fermentation-based bioprocesses based on unsupervised learning (clustering) is presented. As a prior identification of phenotypes and conditional interrelations is desired to control fermentation performance, an automated learning method to output reference phenotypes (defined as vector of biomass-specific rates) was developed and the necessary computing process and parameters were assessed. For its demonstration, time series data of 90 cultivations were used which feature a broad spectrum of solventogenic and acidogenic phenotypes, while 14 clusters of phenotypic manifestations were identified.

Improvement of glycine biosynthesis from one-carbon compounds and ammonia catalyzed by the glycine cleavage system in vitro.

Glycine cleavage system (GCS) plays a central role in one-carbon (C1) metabolism and receives increasing interest as a core part of the recently proposed reductive glycine pathway (rGlyP) for assimilation of CO and formate. Despite decades of research, GCS has not yet been well understood and kinetic data are barely available. This is to a large degree because of the complexity of GCS, which is composed of four proteins (H, T, P, and L) and catalyzes reactions involving different substrates and cofactors.

A novel downstream process for highly pure 1,3-propanediol from an efficient fed-batch fermentation of raw glycerol by .

An efficient downstream process without prior desalination was developed for recovering 1,3-propanediol (1,3-PDO) with high purity and yield from broth of a highly productive fed-batch fermentation of raw glycerol by . After removal of biomass and proteins by ultrafiltration, and concentration by water evaporation, 1,3-PDO was directly recovered from the broth by vacuum distillation with continuous addition and regeneration of glycerol as a supporting agent. Inorganic salts in the fermentation broth were crystallized but well suspended by a continuous flow of glycerol during the distillation process, which prevented salt precipitation and decline of heat transfer.

Dynamic energy correlation analysis of aspartokinase III and alteration of allosteric regulation by manipulating energy transduction pathways.

Conformational change associated with allosteric regulation in a protein is ultimately driven by energy transformation. However, little is known about the latter process. In this work, we combined steered molecular dynamics simulations and sequence conservation analysis to investigate the conformational changes and energy transformation in the allosteric enzyme aspartokinase III (AK III) from .

Tunable Protein Hydrogels: Present State and Emerging Development.

In recent years, protein and peptide-based hydrogels have received great attention for applications in biomedicine. Compared to hydrogels based on synthetic materials, they have the decisive advantages of being biological origin, providing cells with a more in vivo-like microenvironment and possessing potential biological activity. Empowered by the steadily deepened understanding of the sequence-structure-function relationship of natural proteins and the rapid development of molecular-biological tools for accurate protein sequence editing, researchers have developed a series of recombinant proteins as building blocks and responsive blocks to design novel functional hydrogels.

An Aldolase-Based New Pathway for Bioconversion of Formaldehyde and Ethanol into 1,3-Propanediol in .

Formaldehyde (HCHO) is a reactive one-carbon compound that is interesting for biosynthesis. The assimilation of HCHO depends on the catalysis of aldolase. Here, we present a novel synthetic pathway in to convert HCHO and ethanol into 1,3-propanediol (PDO) using a deoxyribose-5-phosphate aldolase (DERA).